DNA-based dual fluorescence signals on and ratiometric mercury sensing in fetal calf serum with simultaneous excitation

Compared to the single emission intensity based sensors, the ratiometric fluorescence assay has achieved improved sensitivity and accuracy by providing built-in correction. However, the dual signals on and ratiometric methods have rarely been reported. Here, a novel strategy for constructing dual fluorescence signals on and ratiometric mercury sensing system is proposed using two kinds of fluorescence dyes (4′,6-diamidino-2-phenylindole (DAPI) and N-methylmesoporphyrin IX (NMM)) and two DNA sequences with both thymine (T)-rich and guanine (G)-rich at two ends. In the presence of Hg2+, T-rich segments cooperatively coordinate with Hg2+ due to the specific and strong affinity between T-T mismatched base pairs and Hg2+, which causes the formation of the split G-quadruplex by two G-rich segments at the other ends. So, both the fluorescent dyes of DAPI and NMM selectively intercalate into dsDNA and G-quadruplex DNA separately, and their fluorescence at 455 nm and 610 nm were enhanced accordingly. Under the experimental conditions employed here, the ratiometric signal was proportional to the Hg2+ concentrations from 0.05 to 3 μM. The limit of detection (LOD) was estimated to be 8 nM based on 3s/m rule. The Hg2+ concentration in FCS has been successfully determined, further demonstrating the potential utility of the novel system.